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Radical prostatectomy for prostate cancer can not only induce cavernous nerve injury (CNI), but also causes cavernous hypoxia and cavernous structural changes, which lead to a poor response to phosphodiesterase 5 inhibitors. To investigate the therapeutic effect of oral administration of LM11A-31, a small molecule p75 neurotrophin receptor (p75NTR) ligand and proNGF antagonist, in a mouse model of bilateral CNI, which mimics nerve injury–induced erectile dysfunction after radical prostatectomy. 8-week-old male C57BL/6 mice were divided into sham operation and CNI groups. Each group was divided into 2 subgroups: phosphate-buffered saline and LM11A-31 (50 mg/kg/day) being administered once daily starting 3 days before CNI via oral gavage. 2 weeks after CNI, we measured erectile function by electrical stimulation of the bilateral cavernous nerve. The penis was harvested for histologic examination and Western blot analysis. The major pelvic ganglia was harvested and cultured for assays of ex vivo neurite outgrowth. Intracavernous pressure, neurovascular regeneration in the penis, in vivo or ex vivo functional evaluation, and cell survival signaling were measured. Erectile function was decreased in the CNI group (44% of the sham operation group), while administration of LM11A-31 led to a significant improvement of erectile function (70% of the sham operation group) in association with increased neurovascular content, including cavernous endothelial cells, pericytes, and neuronal processes. Immunohistochemical and Western blot analyses showed significantly increased p75NTR expression in the dorsal nerve of CNI mice, which was attenuated by LM11A-31 treatment. Protein expression of active PI3K, AKT, and endothelial nitric oxide synthase was increased, and cell death and c-Jun N-terminal kinase signaling was significantly attenuated after LM11A-31 treatment. Furthermore, LM11A-31 promoted neurite sprouting in cultured major pelvic ganglia after lipopolysaccharide exposure. LM11A-31 may be used as a strategy to treat erectile dysfunction after radical prostatectomy or in men with neurovascular diseases. Unlike biological therapeutics, such as proteins, gene therapies, or stem cells, the clinical application of LM11A-31 would likely be relatively less complex and low cost. Our study has some limitations. Future studies will assess the optimal dosing and duration of the compound. Given its plasma half-life of approximately 1 hour, it is possible that dosing more than once per day will provide added efficacy. Specific inhibition of the proNGF-p75NTR degenerative signaling via oral administration of LM11A-31 represents a novel therapeutic strategy for erectile dysfunction induced by nerve injury. Yin GN, Ock J, Limanjaya A, et al. Oral Administration of the p75 Neurotrophin Receptor Modulator, LM11A-31, Improves Erectile Function in a Mouse Model of Cavernous Nerve Injury. J Sex Med 2021;18:17–28. [ABSTRACT FROM AUTHOR]

Extracellular vesicle (EV)–mimetic nanovesicles (NVs) from embryonic stem cells have been observed to stimulate neurovascular regeneration in the streptozotocin-induced diabetic mouse. Pericytes play important roles in maintaining penile erection, yet no previous studies have explored the effects of pericyte-derived NVs (PC-NVs) in neurovascular regeneration in the context of erectile dysfunction. To investigate the potential effect of PC-NVs in neurovascular regeneration. PC-NVs were isolated from mouse cavernous pericytes, and neurovascular regeneration was evaluated in an in vitro study. Twelve-week-old C57BL/6J mice were used to prepare cavernous nerve injury model. Erectile function evaluation, histologic examination of the penis, and Western blots were assessed 2 weeks after model creation and PC-NVs treatment. The main outcomes of this study are PC-NVs characterization, intracavernous pressure, neurovascular regeneration in the penis, and in vitro functional evaluation. The PC-NVs were extracted and characterized by cryotransmission electron microscopy and EV-positive (Alix, TSG101, CD81) and EV-negative (GM130) markers. In the in vivo studies, PC-NVs successfully improved erectile function in cavernous nerve injury mice (∼82% of control values). Immunofluorescence staining showed significant increases in pericytes, endothelial cell, and neuronal contents. In the in vitro studies, PC-NVs significantly increased mouse cavernous endothelial cells tube formation, Schwann cell migration, and dorsal root ganglion and major pelvic ganglion neurite sprouting. Finally, Western blot analysis revealed that PC-NVs upregulated cell survival signaling (Akt and eNOS) and induced the expression of neurotrophic factors (brain-derived neurotrophic factor, neurotrophin-3, and nerve growth factor). PC-NVs may be used as a strategy to treat erectile dysfunction after radical prostatectomy or in men with neurovascular diseases. We evaluated the effect of PC-NVs in vitro and in a mouse nerve injury model, cavernous nerve injury. Additional studies are necessary to determine the detailed mechanisms of neurovascular improvement. Further study is needed to test whether PC-NVs are also effective when given weeks or months after nerve injury. PC-NVs significantly improved erectile function by enhancing neurovascular regeneration. Local treatment with PC-NVs may represent a promising therapeutic strategy for the treatment of neurovascular diseases. Yin GN, Park S-H, Ock J, et al. Pericyte-Derived Extracellular Vesicle–Mimetic Nanovesicles Restore Erectile Function by Enhancing Neurovascular Regeneration in a Mouse Model of Cavernous Nerve Injury. J Sex Med 2020;17:2118–2128. [ABSTRACT FROM AUTHOR]

Abstract Introduction Penile neurovascular dysfunction is a major cause of erectile dysfunction (ED) in diabetic patients, which causes poor response to oral phosphodiesterase-5 inhibitors. Nerve growth factor precursor (proNGF) and its p75 neurotrophin receptor (p75NTR) have been known to be involved in microvascular complications and neurodegeneration. Aim To examine the role of proNGF and its receptor p75NTR signaling pathway in diabetic ED, and to determine the effectiveness of proNGF neutralizing antibody (proNGF-Ab) in restoring erectile function in streptozotocin (STZ)-induced diabetic mice. Methods Diabetes mellitus was induced by intraperitoneal injection of STZ (50 mg/kg) into 8-week-old C57BL/6 male mice for 5 consecutive days. At 8 weeks after the induction of diabetes mellitus, the animals were distributed into 3 groups: controls and STZ-induced diabetic mice receiving 2 intracavernous injections of either saline (days −3 and 0; 20 μL) or proNGF-Ab (days −3 and 0; 20 μg in 20 μL of saline). We also examined the effect of proNGF-Ab or p75NTR small interfering RNA in primary cultured mouse cavernous endothelial cells, pericytes, and major pelvic ganglion. Main Outcome Measures Erectile function was measured by electrical stimulation of the cavernous nerve at 2 weeks after treatment, and the penis was then harvested for histologic and biochemical studies. Results The cavernous expression of proNGF and p75NTR was upregulated under diabetic conditions. Intracavernous injection of proNGF-Ab successfully restored erectile function in diabetic mice, which reach 93–96% of control values. ProNGF-Ab significantly restored cavernous endothelial cell, pericyte, and neuronal cell content, and increased endothelial cell-to-cell junction proteins in the diabetic mice. Under the high-glucose condition, proNGF-Ab or p75NTR small interfering RNA promoted tube formation in mouse cavernous endothelial cells and pericytes, decreased apoptosis of endothelial cells and pericytes, and enhanced neurite sprouting from major pelvic ganglion. Clinical Implications The ProNGF/p75NTR pathway will be a new therapeutic target for diabetic ED. Strength & Limitations This is the first study demonstrating the efficacy of the inhibition of proNGF/p75NTR pathway in diabetic ED. Further studies are needed to test whether a different dosing of proNGF-Ab would induce more durable erectile function recovery. Conclusion Our findings suggest that inhibition of the proNGF/p75NTR signaling pathway is a promising therapeutic strategy for diabetic ED. Nguyen NM, Song K-M, Choi M-J, et al. Inhibition of proNGF and p75NTR Pathway Restores Erectile Function Through Dual Angiogenic and Neurotrophic Effects in the Diabetic Mouse. J Sex Med 2019;16:351–364. [ABSTRACT FROM AUTHOR]

Introduction Diabetic erectile dysfunction is a disease mostly of vascular origin and men with diabetic erectile dysfunction respond poorly to oral phosphodiesterase-5 inhibitors. Hepatocyte growth factor (HGF) is a pleiotropic factor that plays an essential role in the regulation of cell proliferation, survival, and angiogenesis. Aim To determine the effectiveness of recombinant human (rh)-HGF in restoring erectile function in diabetic mice. Methods Four groups of mice were used: control non-diabetic mice and streptozotocin-induced diabetic mice receiving two successive intracavernous injections of phosphate buffered saline (days −3 and 0), a single intracavernous injection of rh-HGF (day 0), or two successive intracavernous injections of rh-HGF (days −3 and 0). We also examined the effect of rh-HGF in primary cultured mouse cavernous endothelial cells and in major pelvic ganglion culture in vitro, which was incubated under a normal-glucose (5 mmol/L) or a high-glucose (30 mmol/L) condition. Main Outcome Measures Two weeks after treatment, we measured erectile function by electrical stimulation of the cavernous nerve and the penis was harvested for histologic studies. Results Repeated intracavernous injections of rh-HGF protein induced significant restoration of erectile function in diabetic mice (89–100% of control values), whereas a single intracavernous injection of rh-HGF protein elicited modest improvement. Rh-HGF significantly induced phosphorylation of its receptor c-Met, increased the content of endothelial cells and smooth muscle cells, and decreased the generation of reactive oxygen species (superoxide anion and peroxynitrite) and extravasation of oxidized low-density lipoprotein in diabetic mice. Under the high-glucose condition, rh-HGF protein also promoted tube formation in mouse cavernous endothelial cells and enhanced neurite sprouting in major pelvic ganglion culture in vitro. Conclusion The dual angiogenic and neurotrophic effects of HGF could open a new avenue through which diabetic erectile dysfunction can be treated. [ABSTRACT FROM AUTHOR]

Introduction Erectile dysfunction ( ED) is a major complication of radical prostatectomy. Men with radical prostatectomy-induced ED respond less positively to oral phosphodiesterase-5 inhibitors. Aim The study aims to examine whether and how stromal vascular fraction ( SVF) restores erectile function in mice with cavernous nerve injury ( CNI). Methods Twelve-week-old male C57BL/6J mice were used and the animals were distributed into five groups: sham operation group and CNI group receiving a single intracavernous injection of phosphate-buffered saline ( PBS) or SVF (1 × 104, 1 × 105, or 3 × 105 cells/20 μL, respectively). SVF was isolated from epididymal adipose tissues of green fluorescence protein transgenic mice. Main Outcome Measures Two weeks after injection, erectile function was measured by cavernous nerve stimulation. The penis was stained with antibodies to platelet/endothelial cell adhesion molecule-1, phosphohistone H3, and phosphorylated endothelial nitric oxide synthase (phospho- eNOS). We also performed Western blot for angiopoietin-1 ( Ang-1), vascular endothelial growth factor- A, hepatocyte growth factor, phospho- eNOS, and eNOS in the corpus cavernosum tissue. Results Local delivery of SVF restored erectile function in a dose-dependent manner in CNI mice. The highest erectile response was noted at a dose of 3 × 105 cells, for which the response was comparable with that in the sham operation group. Local delivery of SVF significantly increased the expression of angiogenic factor proteins and induced cavernous endothelial cell proliferation and eNOS phosphorylation compared with that in the PBS-treated CNI group. SVF-induced promotion of cavernous angiogenesis and erectile function was diminished in the presence of soluble antibody to Tie2, a receptor tyrosine kinase of Ang-1. Conclusion Secretion of angiogenic factors from SVF is an important mechanism by which SVF induces cavernous endothelial regeneration and restores erectile function. These findings suggest that cavernous endothelial regeneration by using SVF may represent a promising treatment strategy for radical prostatectomy-induced ED. Song K-M, Jin H-R, Park J-M, Choi MJ, Kwon M-H, Kwon K-D, Batbold D, Yin GN, Kim WJ, Koh GY, Ryu J-K, and Suh J-K. Intracavernous delivery of stromal vascular fraction restores erectile function through production of angiogenic factors in a mouse model of cavernous nerve injury. J Sex Med 2014;11:1962-1973. [ABSTRACT FROM AUTHOR]

Introduction Recently, much attention has focused on stem cell therapy; bone marrow-derived stem cells ( BMSCs) are one of the most studied mesenchymal stem cells used in the field of erectile dysfunction ( ED). However, a major limitation for the clinical application of stem cell therapy is the heterogeneous nature of the isolated cells, which may cause different treatment outcomes. Aim We investigated the effectiveness of mouse clonal BMSCs obtained from a single colony by using subfractionation culturing method ( SCM) for erectile function in a mouse model of cavernous nerve injury ( CNI). Methods Twelve-week-old C57 BL/6 J mice were divided into four groups: sham operation group, bilateral CNI group receiving a single intracavernous ( IC) injection of phosphate-buffered saline (20 μL) or clonal BMSCs (3 × 105 cells/20 μL), and receiving a single intraperitoneal ( IP) injection of clonal BMSCs (3 × 105 cells/20 μL). Main Outcome Measures The clonal BMSC line was analyzed for cell-surface epitopes by using fluorescence-activated cell sorting and for differentiation potential. Two weeks after CNI and treatment, erectile function was measured by electrically stimulating the cavernous nerve. The penis was harvested for histologic examinations and Western blot analysis. Results Clonal BMSCs expressed cell surface markers for mesenchymal stem cells and were capable of differentiating into several lineages, including adipogenic, osteogenic, and chondrogenic cells. Both IC and IP injections of clonal BMSCs significantly restored cavernous endothelial and smooth muscle content, and penile nNOS and neurofilament content in CNI mice. IC injection of clonal BMSCs induced significant recovery of erectile function, which reached 90-100% of the sham control values, whereas IP injection of clonal BMSCs partially restored erectile function. Conclusion We established a homogeneous population of mouse clonal BMSCs using SCM; clonal BMSCs successfully restored erectile function in CNI mice. The homogeneous nature of clonal mesenchymal stem cells may allow their clinical applications. Ryu J-K, Kim D-H, Song KM, Yi TG, Suh J-K, and Song SU. Intracavernous delivery of clonal mesenchymal stem cells restores erectile function in a mouse model of cavernous nerve injury. J Sex Med 2014;11:411-423. [ABSTRACT FROM AUTHOR]

Introduction Men with erectile dysfunction ( ED) respond poorly to oral phosphodiesterase-5 inhibitors following radical prostatectomy. Recent studies have reported that up-regulation of transforming growth factor-β1 ( TGF-β1) and activation of the Smad signaling pathway play important roles in cavernous fibrosis and in the deterioration of erectile function in a mouse model of cavernous nerve injury ( CNI) and in patients with spinal cord injury. The mothers against decapentaplegic homolog 7 ( Smad7) is known to inhibit the phosphorylation of Smad2 and Smad3. Aim To investigate the effectiveness of adenoviruses encoding Smad7 gene ( Ad- Smad7) on erectile function in a mouse model of CNI. Methods Twelve-week-old C57 BL/6 J mice were used and distributed into 7 groups: sham operation group, untreated CNI group, and CNI groups receiving a single intracavernous injection of adenovirus encoding Lac Z (1 × 108 virus particles [vp]/20 μL) or adenovirus encoding Smad7 ( Ad- Smad7; 1 × 107, 1 × 108, 2 × 108, or 1 × 109 vp/20 μL). Main Outcome Measures Two weeks after bilateral cavernous nerve crushing and treatment, erectile function was measured by electrical stimulation of the cavernous nerve. The penis was harvested for histologic examinations and Western blot analysis. Results The highest erectile response was noted in CNI mice treated with Ad- Smad7 at a dose of 1 × 108 vp, which reached up to 82-85% of sham control values. Local delivery of Ad- Smad7 significantly decreased endothelial cell apoptosis and the production of extracellular matrix proteins, including plasminogen activator inhibitor-1, fibronectin, collagen I, and collagen IV, and induced endothelial nitric oxide synthase phosphorylation in the corpus cavernosum tissue of CNI mice. Conclusion The adenovirus-mediated gene transfer of Smad7 successfully restored erectile function by enhancing endothelial cell function and through antifibrotic effects. These findings suggest that inhibition of the TGF-β signaling pathway by use of Smad7 may represent a promising therapeutic strategy for ED induced by radical prostatectomy. Song KM, Chung J-S, Choi MJ, Jin H-R, Yin GN, Kwon M-H, Park J-M, Kim WJ, Lee S-J, Kim S-J, Ryu J-K, and Suh J-K. Effectiveness of intracavernous delivery of adenovirus encoding Smad7 gene on erectile function in a mouse model of cavernous nerve injury. J Sex Med 2014;11:51-63. [ABSTRACT FROM AUTHOR]

Introduction Much attention has recently been focused on therapeutic angiogenesis as a treatment for erectile dysfunction ( ED). The apelin and apelin receptor ( APJ) system is known to cause endothelium-dependent vasodilatation and to be involved in angiogenesis. Aim To examine the differential expression of apelin and APJ in animal models of vasculogenic ED and to determine whether and how enhancement of apelin- APJ signaling restores erectile function in hypercholesterolemic mice. Methods Acute cavernous ischemia was induced in C57 BL/6 J mice by bilateral occlusion of internal iliac arteries, and chronic vasculogenic ED was induced by feeding a high-cholesterol diet or by intraperitoneal injection of streptozotocin. Main Outcome Measures Messenger RNA ( mRNA) levels of apelin and APJ were determined in cavernous tissue of each vasculogenic ED model by semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR). We evaluated erectile function by electrical stimulation of the cavernous nerve in hypercholesterolemic mice 1, 3, 7, and 14 days after a single intracavernous injection of apelin protein (5 μg/20 μL). The penis was harvested for histologic examinations and Western blot analysis. Results The cavernous mRNA expression of apelin and APJ was up-regulated in acute ischemia model and down-regulated in chronic vasculogenic ED models. A significant restoration of erectile function was noted 1 day after injection of apelin protein into the penis of hypercholesterolemic mice; however, erectile function returned to baseline values thereafter. The beneficial effects of apelin on erectile function resulted mainly from an activation of endothelial nitric oxide synthase and increase in nitric oxide bioavailability through reduction in reactive oxygen species-mediated endothelial apoptosis rather than through direct endothelial cell proliferation. Conclusion These findings suggest that apelin- APJ signaling is a potential therapeutic target in the treatment of vasculogenic ED. Further studies are needed to develop a potent agonist for APJ and to determine the role of repeated dosing of apelin on long-term recovery of erectile function. Kwon M-H, Tuvshintur B, Kim WJ, Jin H-R, Yin GN, Song K-M, Choi MJ, Kwon K-D, Batbold D, Ryu J-K, and Suh J-K. Expression of the apelin-APJ pathway and effects on erectile function in a mouse model of vasculogenic erectile dysfunction. J Sex Med 2013;10:2928-2941. [ABSTRACT FROM AUTHOR]

Introduction Erectile dysfunction ( ED) is a highly prevalent complication of diabetes, and the severity of endothelial dysfunction is one of the most important factors in reduced responsiveness to oral phosphodiesterase type 5 inhibitors. Aim To study the effects of human angiopoietin-4 ( Ang-4) protein on erectile function in diabetic mice. Methods Diabetes was induced by intraperitoneal injection of streptozotocin into 8-week-old C57BL/ 6J male mice. At 8 weeks after the induction of diabetes, the animals were divided into four groups: control nondiabetic mice and diabetic mice receiving two successive intracavernous injections of phosphate buffered saline (days −3 and 0), a single intracavernous injection of Ang-4 protein (day 0), or two successive intracavernous injections of Ang-4 protein (days −3 and 0). Main Outcome Measures One week after treatment, we measured erectile function by electrical stimulation of the cavernous nerve. The penis was harvested and stained with hydroethidine or antibodies to Ang-4, platelet/endothelial cell adhesion molecule-1, and phosphorylated endothelial nitric oxide synthase ( eNOS). We also determined the differential expression of Ang-4 in cavernous tissue in the control and diabetic mice. The effect of Ang-4 protein on the phosphorylation of Tie-2, Akt, and eNOS was determined in human umbilical vein endothelial cells ( HUVECs) by Western blot. Results The cavernous expression of Ang-4 was downregulated in diabetic mice; Ang-4 was mainly expressed in endothelial cells. Local delivery of Ang-4 protein significantly increased cavernous endothelial content, induced eNOS phosphorylation, and decreased the generation of superoxide anion and apoptosis in diabetic mice. Ang-4 protein strongly increased the phosphorylation of Tie-2, Akt, and eNOS in HUVECs. Repeated intracavernous injections of Ang-4 induced significant restoration of erectile function in diabetic mice (87% of control values), whereas a single intracavernous injection of Ang-4 protein elicited modest improvement. Conclusions Cavernous endothelial regeneration by use of Ang-4 protein may have potential for the treatment of vascular disease-induced ED, such as diabetic ED. Kwon M-H, Ryu J-K, Kim WJ, Jin H-R, Song K-M, Kwon K-D, Batbold D, Yin GN, Koh GY, and Suh J-K. Effect of intracavernous administration of angiopoietin-4 on erectile function in the streptozotocin-induced diabetic mouse. J Sex Med 2013;10:2912-2927. [ABSTRACT FROM AUTHOR]

Introduction. Radical prostatectomy for prostate cancer can not only induce cavernous nerve injury ( CNI) but also result in structural changes in the cavernous tissues. Nerve injury-induced protein 1, Ninjurin-1 ( Ninj1), is known to be involved in neuroinflammatory processes and to be related to vascular regression during the embryonic period. Aim. The study aims to determine whether and how Ninj1 neutralizing antibody ( Ninj1- Ab) restores erectile function in mice with CNI. Methods. Twelve-week-old C57 BL/6 J mice were used and distributed into four groups: sham operation group and CNI groups receiving a single intracavernous injection of immunoglobulin G ( IgG) control antibody, low-dose Ninj1- Ab (1.0 μg/20 μ L), or high-dose Ninj1- Ab (2.5 μg/20 μ L). Main Outcome Measures. One week after bilateral cavernous nerve crush, erectile function was measured by electrical stimulation of the cavernous nerve. The penis was harvested for histologic examinations and Western blot analysis. Results. The cavernous expression of Ninj1 protein was upregulated up to 7 days after CNI and returned to baseline levels thereafter. Local delivery of Ninj1- Ab significantly increased penile neuronal nitric oxide synthase and neurofilament contents, induced cavernous endothelial proliferation and phosphorylation of Akt and endothelial nitric oxide synthase, and decreased endothelial cell apoptosis in the CNI mice by upregulating angiopoietin-1 and downregulating angiopoietin-2. High-dose Ninj1- Ab induced profound restoration of erectile function in the CNI mice (91% of sham control values), whereas low-dose Ninj1- Ab elicited partial improvement. Conclusion. The dual neurotrophic and angiogenic effects of Ninj1 blockade may provide a good opportunity for treating erectile dysfunction resulting from radical prostatectomy. [ABSTRACT FROM AUTHOR]

ABSTRACT Introduction. Men with diabetic erectile dysfunction (ED) often have severe endothelial dysfunction and respond poorly to oral phosphodiesterase-5 inhibitors. Aim. To examine whether and how freshly isolated stromal vascular fraction (SVF) promotes cavernous endothelial regeneration and restores erectile function in diabetic animals. Methods. Eight-week-old C57BL/6J mice were used. Diabetes was induced by intraperitoneal injection of streptozotocin. SVF was isolated from epididymal adipose tissues of green fluorescence protein transgenic mice. At 8 weeks after the induction of diabetes, the animals were divided into six groups: controls, diabetic mice, and diabetic mice treated with a single intracavernous injection of phosphate-buffered saline (PBS) or SVF (1 × 104 cells, 1 × 105 cells, or 2 × 105 cells/20 µL, respectively). Main Outcome Measures. Two weeks later, erectile function was measured by cavernous nerve stimulation. The penis was stained with antibodies to CD31, CD34, phosphohistone H3, phospho-endothelial nitric oxide synthase (eNOS), and vascular endothelial growth factor-A (VEGF-A). We also performed Western blot for phospho-eNOS and eNOS, and determined cyclic guanosine monophosphate (cGMP) concentration in the corpus cavernosum tissue. Results. Significant improvement in erectile function was noted in diabetic mice treated with SVF at concentrations of 1 × 105 and 2 × 105 cells, which reached up to 82% of the control values. Local delivery of SVF significantly increased cavernous endothelial cell proliferation, eNOS phosphorylation, and cGMP expression compared with that in the untreated group and the PBS-treated diabetic group. Intracavernous injection of SVF increased cavernous VEGF-A expression and induced recruitment of CD34(+)CD31(−) progenitor cells. Some SVF underwent differentiation into cavernous endothelial cells. SVF-induced promotion of cavernous angiogenesis and erectile function was abolished in the presence of VEGF-Trap, a soluble VEGF-A neutralizing antibody. Conclusion. The results support the concept of cavernous endothelial regeneration by use of SVF as a curative therapy for diabetic ED. Ryu J-K, Tumurbaatar M, Jin H-R, Kim WJ, Kwon M-H, Piao S, Choi MJ, Yin GN, Song K-M, Kang Y-J, Koh YJ, Koh GY, and Suh J-K. Intracavernous delivery of freshly isolated stromal vascular fraction rescues erectile function by enhancing endothelial regeneration in the streptozotocin-induced diabetic mouse. J Sex Med 2012;9:3051-3065. [ABSTRACT FROM AUTHOR]

ABSTRACT Introduction. A proper cavernous endothelial cell culture system would be advantageous for the study of the pathophysiologic mechanisms involved in endothelial dysfunction and erectile dysfunction (ED). Aim. To establish a nonenzymatic technique, which we termed the 'Matrigel-based sprouting endothelial cell culture system,' for the isolation of mouse cavernous endothelial cells (MCECs) and an in vitro model that mimics in vivo situation for diabetes-induced ED. Methods. For primary MCEC culture, mouse cavernous tissue was implanted into Matrigel and sprouting cells from the tissue were subcultivated. To establish an in vitro model for diabetes-induced ED, the primary cultured MCECs were exposed to a normal-glucose (5 mmoL) or a high-glucose (30 mmoL) condition for 48 hours. Main Outcome Measures. The purity of isolated cells was determined by fluorescence-activated cell sorting analysis. MCECs incubated under the normal- or the high-glucose condition were used for Western blot, cyclic guanosine monophosphate (cGMP) quantification, and in vitro angiogenesis assay. Results. We could consistently isolate high-purity MCECs (about 97%) with the Matrigel-based sprouting endothelial cell culture system. MCECs were subcultured up to the fifth passage and no significant changes were noted in endothelial cell morphology or purity. The phosphorylation of Akt and eNOS and the cGMP concentration were significantly lower in MCECs exposed to high glucose than in those exposed to normal glucose. MCECs exposed to the normal-glucose condition formed well-organized capillary-like structures, whereas derangements in tube formation were noted in MCECs exposed to high glucose. The protein expression of transforming growth factor-β1 (TGF-β1) and phospho-Smad2 was significantly increased by exposure to high glucose. Conclusion. The Matrigel-based sprouting endothelial cell culture system is a simple, technically feasible, and reproducible technique for isolating pure cavernous endothelial cells in mice. An in vitro model for diabetic ED will be a valuable tool for evaluating the angiogenic potential of novel endogenous or synthetic modulators. Yin GN, Ryu J-K, Kwon M-H, Shin SH, Jin HR, Song K-M, Choi MJ, Kang D-Y, Kim WJ, and Suh J-K. Matrigel-based sprouting endothelial cell culture system from mouse corpus cavernosum is potentially useful for the study of endothelial and erectile dysfunction related to high-glucose exposure. J Sex Med 2012;9:1777-1789. [ABSTRACT FROM AUTHOR]

ABSTRACT Introduction. Transforming growth factor-β1 (TGF-β1) has been identified as an important fibrogenic cytokine associated with Peyronie's disease (PD). Aim. The aim of this study was to study the differential expression of the TGF-β1 and Smad transcription factors in plaque tissue from PD patients and to determine the antifibrotic effect of SKI2162 (SK Chemicals, Seoul, South Korea), a novel small-molecule inhibitor of activin receptor-like kinase 5 (ALK5), a type I receptor of TGF-β, in primary fibroblasts derived from human PD plaque. Methods. Plaque tissue was isolated from five PD patients, and tunica albuginea tissue was obtained from four control patients. Plaque tissues from a patient with PD were used for primary fibroblast culture. Fibroblasts were pretreated with SKI2162 (10 µM) and then stimulated with TGF-β1 (10 ng/mL). Main Outcome Measures. The plaque or tunica albuginea tissue was stained with Masson's trichrome or antibody to TGF-β1, phospho-Smad2 (P-Smad2), and P-Smad3. Protein was extracted from treated fibroblasts for Western blotting, and the membranes were probed with antibody to P-Smad2/Smad2, P-Smad3/Smad3, plasminogen activator inhibitor-1, fibronectin, collagen I, and collagen IV. We also determined the inhibitory effect of SKI2162 on TGF-β1-induced nuclear translocation of Smad2/3 in fibroblasts. Results. The plaque tissue from PD patients showed higher TGF-β1, P-Smad2, and P-Smad3 immunoreactivity than did the tunica albuginea tissue from control patients. SKI2162 not only blocked TGF-β1-induced phosphorylation and nuclear translocation of Smad2 and Smad3, but also inhibited the production of extracellular matrix markers in fibroblasts derived from human PD plaque. Conclusion. In light of the pivotal role of TGF-β and Smads in the pathogenesis of PD, pharmacologic inhibition of ALK5 may represent a novel targeted approach to treating PD. Piao S, Choi MJ, Tumurbaatar M, Kim WJ, Jin H-R, Shin SH, Tuvshintur B, Yin GN, Song JS, Kwon M-H, Lee S-J, Han J-Y, Kim S-J, Ryu J-K, and Suh J-K. Transforming growth factor (TGF)-β type I receptor kinase (ALK5) inhibitor alleviates profibrotic TGF-β1 responses in fibroblasts derived from Peyronie's plaque. J Sex Med 2010;7:3385-3395. [ABSTRACT FROM AUTHOR]

ABSTRACT Introduction. With the advent of genetically engineered mice, it seems important to develop a mouse model of cavernous nerve injury (CNI). Aim. To establish a mouse model of CNI induced either by nerve crushing or by neurectomy and to evaluate time-dependent derangements in penile hemodynamics in vivo and subsequent histologic alterations in the cavernous tissue. Methods. Twelve-week-old C57BL/6J mice were divided into 4 groups (N = 36 per group): control, sham operation, bilateral cavernous nerve crush, and bilateral cavernous neurectomy group. Main Outcome Measures. Three days and 1, 2, 4, 8, and 12 weeks after CNI, erectile function was measured by electrical stimulation of the cavernous nerve. The penis was then harvested and TUNEL was performed. Immunohistochemical analysis was performed assaying for caspase-3, transforming growth factor-β1 (TGF-β1), phospho-Smad2, PECAM-1, factor VIII, and smooth muscle α-actin. The numbers of apoptotic cells and phospho-Smad2-immunopositive cells in endothelial cells or smooth muscle cells were counted. Results. Erectile function was significantly less in the cavernous nerve crushing and neurectomy groups than in the control or sham group. This difference was observed at the earliest time point assayed (day 3) and persisted up to 4 weeks after nerve crushing and to 12 weeks after neurectomy. The apoptotic index peaked at 1 or 2 weeks after CNI and decreased thereafter. Cavernous TGF-β1 and phospho-Smad expression was also increased after CNI. The numbers of apoptotic cells and phospho-Smad2-immunopositive cells in cavernous endothelial cells and smooth muscle cells were significantly greater in the cavernous nerve crush and cavernous neurectomy groups than in the control or sham group. Conclusion. The mouse is a useful model for studying pathophysiologic mechanisms involved in erectile dysfunction after CNI. Early intervention to prevent apoptosis in smooth muscle cells and endothelial cells or to inhibit cavernous tissue fibrosis is required to restore erectile function. Jin H-R, Chung YG, Kim WJ, Zhang LW, Piao S, Tuvshintur B, Yin GN, Shin SH, Tumurbaatar M, Han J-Y, Ryu J-K, and Suh J-K. A mouse model of cavernous nerve injury-induced erectile dysfunction: Functional and morphological characterization of the corpus cavernosum. J Sex Med 2010;7:3351-3364. [ABSTRACT FROM AUTHOR]

Introduction. Mirodenafil is a newly developed oral phosphodiesterase type 5 inhibitor, currently under investigation as a treatment for erectile dysfunction (ED). Aim. We investigated the efficacy and safety of on demand mirodenafil therapy at fixed doses (50 and 100 mg) in Korean men with a broad range of ED. Methods. A multicenter, randomized, double-blind, placebo-controlled, parallel-group, fixed-dose study was conducted with 223 subjects who were randomized to placebo or mirodenafil at fixed doses of 50 or 100 mg for 12 weeks on an “as needed” basis. Main Outcome Measures. Primary efficacy measures were scores on the International Index of Erectile Function (IIEF) Question 3 (Q3) and Question 4 (Q4). Secondary efficacy measures included all domain scores of the IIEF, Sexual Encounter Profile Question 2 (SEP2), Sexual Encounter Profile Question 3 (SEP3), the Global Assessment Question (GAQ), and the Life Satisfaction Checklist (LSC). Safety assessments included laboratory tests, vital signs, physical examination, 12-lead electrocardiogram recordings, and patients’ reporting of adverse events. Results. Mirodenafil 50 and 100 mg groups showed a significantly greater increase in IIEF Q3 ( P = 0.0001, P < 0.0001, respectively) and Q4 scores (both P < 0.0001) at the end point compared with the placebo group. And mirodenafil in both doses significantly improved the scores of all five domains of the IIEF, SEP2, and SEP3 as well as the percentages of patients responding positively to the GAQ compared with the placebo group. As for LSC scores, the two mirodenafil groups showed significantly greater improvements in items regarding life as a whole, sexual life, and partner relationship than the placebo group. Most treatment-associated adverse events were of mild intensity, resolving spontaneously. Conclusions. Mirodenafil, in doses of 50 or 100 mg, significantly improved erectile function and were well tolerated in a representative population of Korean men with broad-spectrum ED of various etiologies and severities. Paick J-S, Ahn TY, Choi HK, Chung W-S, Kim JJ, Kim SC, Kim SW, Lee SW, Min KS, Moon KH, Park JK, Park K, Park NC, Suh J-K, Yang DY, and Jung H-G. Efficacy and safety of mirodenafil, a new oral phosphodiesterase type 5 inhibitor, for treatment of erectile dysfunction. J Sex Med 2008;5:2672–2680. [ABSTRACT FROM AUTHOR]